The rational development of new antineoplastic agents directed against tubulin, a protein critical for cell division, requires greater understanding of the interactions between the polypeptide subunits of tubulin, its two tightly bound guanine nucleotides, and drugs which inhibit mitosis. Substitution of ATP and ATP analogs for GTP in glycerol-induced polymerization was demonstrated. This will permit structure-function studies of nucleotide interactions at the exchangeable site with more readily available ATP analogs. Degradation of the nonexchangeable GTP of tubulin by phosphofructokinase was observed, representing the first approach to this nucleotide which does not require the denaturation of the protein. Studies on the interactions of a large group of antimitotic agents with tubulin continued. The classes of compounds examined were combretastatin congeners; derivatives of 6-benzyl-1,3-benzodioxole and related compounds; derivatives of 5,6-diarylpyridazin-3-one; 2,4-dichlorobenzylthiocyanate; and alkyl carbamates of aromatic amines. A microtubule-associated protein which causes the formation of microtubule bundles was purified to homogeneity. Microtubule-associated protein(s) with nucleoside monophosphate kinase activity were resolved into components with different substrate specificity, and one component was purified to homogeneity. Maytansine was found to inhibit entry and exit of GDP and GTP at the exchangeable nucleotide binding site. Efforts to separate the Alpha and Beta subunits of tubulin continued, and efforts to prepare GTP analogs derivatized at the Gamma-phosphate were initiated.